The striking increase in atherosclerosis in diabetes suggests that factors directly consequent to hyperglycemia impact on the response of the blood vessel wall to superimposed biochemical and metabolic stresses, such as hyperlipidemia. Sustained vascular accumulation of ligands for Receptor for AGE (RAGE), the products of nonenzymatic glycation and oxidation of proteins/lipids, the Advanced Glycation Endproducts (AGEs); and S100/calgranulins; a family of proinflammatory cytokines, provides one such mechanism for acceleration of diabetic atherosclerosis. Pharmacological blockade of RAGE, or transgenic mice in whom RAGE signalling is disabled, secondary to cell-specific introduction of a transgene in which the cytosolic tail is deleted, suppressed acceleration of diabetic atherosclerosis in mice deficient in apolipoprotein (ape) E (0). Nondiabetic homozygous RAGE (0) mice in the ape E (0) background displayed decreased atherosclerosis compared to mice solely deficient in ape E (0). Substantial progress in the first years of the Program Project has been made in identifying the signal transduction cascades recruited by ligand engagement of RAGE. Recent studies in a yeast two-hybrid assay indicated that the cytosolic domain of RAGE interacts with a newly-identified member of the diaphanous family of proteins. Diaphanous molecules, first identified in Drosophila, are critical intracellular "bridges" of cell signalling and modulation of the actin cytoskeleton in mammalian cells, as specific domains of these molecules engage effectors such as the Rho GTPases, thereby impacting on downstream cascades, such as Nuclear Factor (NF)-kB; and polypeptides linked to cellular motility. We hypothesize that RAGE-dependent mechanisms trigger activation of signalling pathways linked to vascular inflammation and cellular motility. Here, we propose to rigorously test the role of RAGE in atherosclerosis using RAGE (0) mice and transgenic mice in whom RAGE signalling is disabled in endothelial cells, smooth muscle cells and cells of mononuclear phagocyte lineage, and to determine the critical domains of RAGE and diaphanous-like protein that mediate their interactions, and, thereby, modulation of cellular motility and signalling. These studies will further our understanding of mechanisms by which RAGE activation is linked to vascular stress, and highlight the impact of this pathway for future therapeutic strategies in atherosclerosis. This Project is closely linked to Project by Yan, as each studies RAGE-dependent vascular stress. This Project shares mouse models and ligand-RAGE reagents with each of the other projects. This Project will utilize all three Cores of the Program Project.